In autosomal dominant polycystic kidney disease (ADPKD), aberrant proliferation of tubular epithelial cells is a major factor in renal cyst formation. We have shown that agonists that elevate intracellular cAMP accelerate the rate of proliferation of ADPKD cells. By contrast, cAMP was found to be anti-mitogenic in normal human kidney (NHK) cells. The mechanism for the phenotypic difference in cAMP mitogenicity between ADPKD and NHK cells is unknown. Recently, we found that cAMP-dependent protein kinase A (PKA) activated the mitogen-activated protein (MAP) kinase pathway involving extracellular signal-regulated kinase (ERK) in ADPKD cells, but not NHK cells. The basis for PKA stimulation of ERK is thought to be dependent on B-Raf abundance. Cells that contain high levels of B-Raf, such as neuronal cells, are stimulated by cAMP, while cells with little B-Raf, such as astrocytes, are inhibited. The proliferative response to cAMP by ADPKD cells is unusual in that renal cells, which are normally inhibited by cAMP, are stimulated to proliferate owing to mutations in polycystins. Recent studies indicate polycystin-1 (PC1) and polycystin-2 (PC2) are partners in the regulation of Ca 2+ entry into cells. We propose that defects in PC1 or PC2 and the loss of polycystin function alter intracellular Ca 2+ and secondarily cause a phenotypic switch in the proliferative response to cAMP. In preliminary experiments, we found that inhibition of Ca 2+ entry with Ca 2+ channel blockers, reducing extracellular Ca 2+, or disrupting PC1/PC2 signaling by overexpressing the C-tail of PC1 switched the phenotype of M1 cells (a mouse cortical collecting duct cell line) from cAMP-inhibitory to cAMP-stimulatory phenotype, mimicking the difference between NHK and ADPKD cells. The M1 cell model provides a method for investigating the molecular mechanisms responsible for the phenotypic switch in the cAMP-dependent proliferation within a particular cell. To elucidate these mechanisms, we will investigate the following aims: 1) Determine the molecular mechanism by which a reduction in the intracellular [Ca 2+] alters cAMP-dependent B-Raf signaling in M1 cells. 2) Determine the mechanism by which the overexpression of the C-terminal fragment of PC1 in M1 cells, a condition thought to disrupt PC1/PC2 function, alters cAMP-dependent B-Raf signaling.